1. Technical Field
This application relates generally to microbial sampling and filtering devices, and more specifically to suction devices for sampling, filtering, recovering and/or concentrating microbial populations.
2. Background
Outbreaks of human enteric diseases caused by food borne E. coli 0157/H7, Salmonella spp., Campylobacter jejumi/coli and Listeria monocytogenes caused an estimated 5,000,000 illnesses and 3,700 deaths in the U.S. in 1993. Costs due to medical and production losses are estimated between $4.7 and $7.5 billion for the same period. Foods are routinely tested for microbial contamination during the process of preparing a food for consumption by consumers. With meats, a main problem is contamination with E. coli bacteria. E. coli bacteria are a type of bacteria which is generally present in the digestive tract and fecal material of animals.
Meat processors are under increasing requirements to ensure that their meat processing systems produce meat which is free of E. coli or other bacteria pathogenic contamination. One program which has been recently introduced is called "Hazards Analysis and Critical Control Point", or HACCP. This program is based on analyzing a system to determine at what critical points increased control would result in a marked improvement of quality or reduction of contamination of the food product. Under this program, food processors are required to analyze their systems and determine at which critical control points increased testing should be utilized. Once these critical control points are identified, increased testing at these critical points should result in better control of the food process, and more assurance of safety of the final food product.
Under the HACCP Program, the USDA will require over 9,000 meat and poultry establishments to conduct more bacterial lab tests than are presently performed. It is also a major goal of microbial lab test protocols on meat and poultry samples to report at least preliminary test results within a few hours of sampling animal carcasses. This ideal goal would likely save millions of dollars annually for the industry, since recalls and holding times of suspect products would be reduced.
In the processing of beef and other meat products, some of the earliest steps are to hang the beef by its hind legs, skin the beef, remove the entrails from the beef, and split the beef in half down the center line. It has been determined that these steps are critical control points and are steps during which there is a heightened possibility of contamination. Since E. coli bacteria exists in the intestinal tract and feces of beef and other animals, the process of skinning the carcass around the anus, and removing the entrails by a worker using a knife, presents a high possibility of the knife becoming contaminated with E. coli bacteria. If a knife thus contaminated is laid down on a surface, other equipment which comes in contact with the same surface can be cross-contaminated. For this reason it is critical to be able to sample the beef carcass after the entrails have been removed and after the skin has been removed, to check for the presence of E. coli bacteria. It is also critical to be able to test other surfaces and equipment, such as the surfaces of knives, table tops or meat grinders to check for the presence of E. coli bacteria.
A variety of non-destructive bacterial sample collection devices are commercially available for sampling large animal carcasses. These include direct agar contact, adhesive contact tape, rinsing, scraping and vacuuming. Vacuuming or aspiration procedures have not been successfully applied to meat animal carcasses. A bacterial, or dust particle vacuuming method, has been used on clean room surfaces. This design would not have application on animal carcasses. The PASS carcass sampler utilizes a sterile spray applied to the carcass surface, followed by collection of the residual liquid by aspiration or pressure. This device is manufactured by pbi of Italy. This sampling device has not been widely accepted in the United States.
The most practical of the current methods for sampling bacteria on larger animal carcasses involve the use of swabs or sponges, with and without templates, which blot, wipe or soak up surface moisture and accompanying microbes from the selected surfaces. These methods can collect a single bacterial colony forming a unit (cfu), if adsorbed during sampling. The principle behind this technique is that for every individual bacterium which was originally on the sterile sponge or cotton swab, each of those individual bacterium will be transferred to the broth and later to the agar in the petri dish. Over a period of about 18 hours at the proper temperature and atmospheric conditions (E. coli are normally aerobic bacteria), each of the individual bacterium (cfu) will have grown by cell division into a colony of bacteria, with each colony on an agar plate being visible to the naked eye. Since these bacteria typically double their population in approximately 20 to 30 minutes under ideal conditions, after 18 hours in nutrient broth or on agar plates, sufficient divisions of the original bacteria will have occurred so that the increased numbers of bacteria may be more readily detected.
This sponge method assumes a consistent increased affinity of the collection device surface over that of the sampled surface during collection. Since bacteria affinity and attachment to carcass surfaces are influenced by surface pH, carcass temperature, texture, hydrophobicity, ionic strength, and surface moisture, actual percentages of total microbes collected may be less than representative.
Many of the bacteria which were on the meat could be trapped within folds of the carcass surface or within rough areas, fat cells or connective tissue and simply not be wiped off onto the sponge. Of those bacteria that do get wiped onto the sponge, many of them may stick to the sponge and not be transferred to the broth solution or the petri dishes during a quick rinse or transfer attempts. The incubation time of 18 hours means that if a beef carcass were severely contaminated, it would have moved along the process for 18 hours and may have cross contaminated other beef carcasses or other cutting utensils or handling machinery.
A further limitation to this method is that moisture saturated sponges may spread pathogens from one location to another during sampling of more than one location on a carcass. Reversing sides of the sponge and sampling the three recommended sites on beef and swine carcasses from least to most likely contaminated, offers a good approach to circumventing this problem. But, if used incorrectly, or in cases of abnormal carcass bacterial distribution, it may contribute to further spreading of pathogens.
Current sampling methods result in routine lab specimens which require several hours of enrichment growth before analysis for bacteria identification. Following enrichment procedures, standard or rapid bacterial detection methods may be used. In an attempt to conduct faster analysis during current collection methods and rapid bacteria detection kits, carcass collection sponges may be rinsed or soaked for short time periods in buffered solutions. Results of these procedures are questionable because bacterial numbers are normally low. False negative lab results are more potentially dangerous to consumers than false positives. Therefore, rapid detection methods are not routinely acceptable with current collection techniques without enrichment steps.
Filtration and centrifugation of dilute liquid samples or broth suspected of containing microbes are commonly used in laboratories to separate debris, and to capture and concentrate bacteria for subsequent identification or other testing. Whole bird rinse solutions could be concentrated with these methods, but large fluid volumes from these animal carcasses are cumbersome for routine lab analysis, especially when a large number of samples are involved.
In recent years there have been great improvements in the detection methods for bacteria. For instance, rapid detection systems for E. coli 0157/H7 and Salmonella are currently available that require only a few hours to complete. Current rapid detection methods do not require culturing, but allow a sample to be analyzed in a period of several hours. The shortcoming with these rapid detection methods is that a fairly concentrated sample is required. Most of these methods are not directly applicable to bird or sponge rinse solution currently used because of the low number of pathogens normally collected from carcasses, plus the dilution effect of the rinses. Time and labor-consuming procedures for enrichment and bacterial concentration are required before using the rapid Elisa, PCR or similar identification tests. Improved rapid sampling and processing methods are needed to efficiently utilize these new bacteria identification techniques. Current non-destructive bacterial sampling methods for large animal carcasses have several drawbacks which may be magnified under mandates and ramifications of the HACCP programs. These methods:
A. allow sampling of a limited carcass area only; PA1 B. result in bacterial solutions too dilute for same day analysis; PA1 C. require extended lab enrichment time; PA1 D. require high labor and time expenditure.
Regardless of the lab procedure, improved bacterial sampling methods, especially for large animal carcasses which cannot utilize whole body rinse techniques, are needed which can facilitate larger surface area sampling without undue increases in labor and material. In addition, new sampling methods would allow the meat and poultry establishments to routinely benefit from the use of recently developed rapid detection methods for E. coli 0157/H7 and other human pathogens.
Accordingly, what is needed is a sampling method and device by which more bacteria are removed from the surface which is sampled. It is a further object that once removed from the surface, more of the collected bacteria which are sampled pass through the sampling system and end up being counted, i.e., improved bacterial recovery. To promote improved recovery, bacteria should be readily collectable from the sampling process and equipment.
It is a further object of this invention to provide a method and apparatus by which bacteria on such surfaces can be detected in a shortened time period.
It is a further object of this invention to provide a sampling method by which large volumes of dilute rinse and bacteria suspensions could provide concentrated bacterial populations, and in a shortened period of time over existing sampling methods and equipment.
Additional objects, advantages and novel features of the invention will be set forth in part in the description as follows, and in part will become apparent to those skilled in the art upon examination of the following, or may be learned by practice of the invention. The objects and advantages of the invention may be realized and attained by means of the instrumentalities and combinations particularly pointed out in the appended claims.